Various cloning vectors and their insert size

S.No.	Vector	Host	Insert size (in Kb)
1.	M13 vectors	E. Coli	3
2.	Plasmid vectors	E. Coli	8
3.	Phagemid vectos	E. Coli	10
4.	λgt10	E. Coli	8
5.	λZAPII	E. Coli	10
6.	λEMBL4	E. Coli	20
7.	λGEM11, λGEM12	E. Coli	25
8.	λ cosmids	E. Coli	35-45
9.	P1 derived artificial chromosomes (PAC)	E. Coli	100-300
10.	Bacterial artificial chromosomes (BAC)	E. Coli	≤300
11.	Yeast artificial chromosomes (YAC)	Sachharomyces cerevisiae	200 - 2000

Problem of quantitative genetics

Problem: Fruit colour of wild Solanum nigrum is controlled by two alleles of a gene (A and a). The frequency of A, p=0.8 and a, q=0.2. In a neighbouring field a tetraploid genotype of S. nigrum was found. After critical examination five distinct genotypes were found; which are AAAA, AAAa, AAaa, Aaaa and aaaa. Following Hardy Weinberg principle and assuming the same allele frequency as that of diploid population, the numbers of phenotypes calculated within a population of 1000 plants are close to one of the following: AAAA : AAAa : AAaa : Aaaa : aaaa

1. 409 : 409 : 154 : 26 : 2

2. 420 : 420 : 140 : 18 : 2

3. 409 : 409 : 144 : 36 : 2

4. 409 : 420 : 144 : 25 : 2                                                      (CSIR June 2016)

Solution:

Frequency of allele A = 0.8 and a = 0.2

	AA	Aa	Aa	aa
AA	AAAA	AAAa	AAAa	AAaa
Aa	AAAa	AAaa	AAaa	Aaaa
Aa	AAAa	AAaa	AAaa	Aaaa
aa	AAaa	Aaaa	Aaaa	aaaa

From above chart phenotypic ratio is,

AAAA : AAAa : AAaa : Aaaa : aaaa = 1 : 4 : 6 : 4 : 1

From Hardy-Weinberg principle:

p⁴ : 4p³q : 6p²q² : 4pq³ :q⁴

for AAAA= p⁴ x 1000 = (0.8)⁴ x 1000

= 409.6 » 409

for AAAa = 4p³q x 1000 = 4x(0.8)³x0.2x1000

= 409.6  » 409

for AAaa = 6p²q² x 1000 = 6x(0.8)²x(0.2)² x1000

= 153.6  » 154

for Aaaa = 4pq³ x 1000 = 4x0.8x(0.2)³ x1000

= 25.6  » 26

for aaaa = q⁴ x 1000 = (0.2)⁴ x1000

= 1.6 » 2

Number of phenotypes are 

AAAA : AAAa : AAaa : Aaaa : aaaa = 409 : 409 : 154 : 26 : 2

Option 1 is correct

Function of Vir genes

The genes responsible for the transfer of the T-DNA region into the host plant are also situated on the Ti plasmid in a region of approximately 40 kb outside the T-DNA, known as the vir (virulence) region. The genes of vir region are not transferred themselves; they only induce the transfer of T-DNA.

These genes have following functions:

Vir genes	Functions
VirA	Encodes acetosyringone (phenolic sensor) receptor protein, functions as autokinase; also activates VirG gene by phosphorylation leading to constitutive expression of all genes
VirB1-B11	Encodes membrane protein, involved in conjugal tube formation through which T-DNA is transport, VirB11 has ATPase activity
VirC	Encodes helicase enzymes, binds to the overdrive region, unwinding of T-DNA
VirD1	Topoisomerase activity- required for T-DNA processing, modulates VirD2 activity
VirD2	VirD2 is an endonuclease- nicks the right border of T-DNA, directs T-DNA through the VirB/VirD4 transfer apparatus, contain nuclear localization sequences (NLS) that promote nuclear uptake of the T-complex
VirD4	Components of transfer apparatus
VirE1	Required for VirE2 export from Agrobacterium
VirE2	Single strand binding protein (SSBP), binds to T-DNA during transfer, forms a membrane channel that transfers the T-strand through the plant plasma membrane, involved in nuclear targeting and passage through nuclear pore complex, contain nuclear localization sequences (NLS), assist nuclear uptake of the T-complex by keeping the T-strand in an unfolded state
VirF	Directs protein coating of T-DNA complex removal by proteasomal machinery
VirG	Master controller DNA binding protein, vir A activates vir G by phosphorylation, vir G dimerises and activates constitutive expression of all vir operons
VirJ	T-DNA export

Some compounds that use as herbicides

S.No.	Chemical compound	How it interferes	Mechanism
1.	DCMU (3-(3,4-dichlorophenyl)-1,1-dimethylurea)	Inhibition of electron transfer	Competes with plasoquinone (QB) for binding site in PSII
2.	FCCP (cyanide-p-trifluoromethoxyphenylhydrazone;)	Uncoupling of phosphorylation from electron transfer	Hydrophobic proton carriers
3.	DNP(2,4-dinitrophenol)		Hydrophobic proton carriers
4.	Valinomycin		K+ ionophore
5.	Thermogenin		In brown adipose tissue, forms proton-conducting pores in inner mitochondrial membrane
6.	Atractyloside	Inhibition of ATP-ADP exchange	Inhibits adenine nucleotide translocase
7.	DCCD(dicyclohexylcarbodiimide)	Inhibition of ATP synthase	Blocks proton flow through Fo and CFo
8.	Aurovertin		Inhibits F1
9.	Oligomycin		inhibits Fo and CFo
10.	Venturicidin
11.	Cyanide	Inhibition of electron transfer	Inhibit cytochrome oxidase
12.	Carbon monoxide
13.	Antimycin A		Blocks electron transfer from cytochrome b to cytochrome c1
14.	Myxothiazol		Prevent electron transfer from Fe-S center to ubiquinone
15.	Rotenone
16.	Amytal
17.	Piericidin A

Source: Lehninger Principles of Biochemistry (Nelson and Cox, 6^th edition)